Molecular Cell Biological Study on Neuronal Apoptosis in Cerebellar Graneul Cells

小脑粒细胞神经元凋亡的分子细胞生物学研究

• 批准号: 10480216
• 负责人: HATANAKA Hiroshi
• 金额: $ 2.75万
• 依托单位: Osaka University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10480216/
• 关键词: Apoptosis; PI3-kinase; SNF-1; Neurotrophin; High potassium; Cereberum; Phosphorylation; Low potassium; SNF1; 小脳顆粒細胞

We analyzed the mechanism underlying cell death using cultured rat cerebellar granule and cerebral cortical neurons, because of their abundance and homogeneity. Cerebellar granule neurons maintained in medium containing 26 mM potassium or in medium (5 mM potassium) with 50 ng/ml BDNF undergo an apoptoic cell death when exposed to 10 μ M LY294002, an inhibitor of PI3-kinase. To investigate the intracellular signaling mechanism of LY294002-induced apoptosis, the activities of Akt and c-Jun N-terminal kinase (JNK) were measured in cells in HKィイD1+ィエD1 (26 mM potassium) or LKィイD1+ィエD1 (5 mM potassium) medium containing BDNF, with or without 10 μ M LY294002. Akt activity decreased following the addition of 10 μ M LY294002. In addition, we found that LY294002 increased the JNK activity, which is known to mediate some types of cell death in PNS neurons. We also observed elevated expression of c-Jun by LY294002 in HKィイD1+ィエD1 +BDNF. These findings demonstrated that apoptosis induced by inhibition of PI3-kinase activity involves suppression of the Akt activity and elevation of the JNK activity in cerebellar granule neurons. Our results suggested that the PI3-kinase-Akt pathway suppresses the activation of JNK and c-Jun expression, and as a result prevents the neuronal cell death in cerebellar granule neurons.

我们使用培养的大鼠小脑颗粒和大脑皮层神经元分析了细胞死亡的机制，因为它们的丰度和同质性维持在含有 26 mM 钾的培养基或含有 50 ng/ml BDNF 的培养基（5 mM 钾）中。暴露于 10 μM PI3 激酶抑制剂 LY294002 时细胞凋亡 探讨细胞内信号传导机制。 LY294002诱导的细胞凋亡，在含有BDNF的HKD1+D1（26 mM钾）或LKD1+D1（5 mM钾）培养基中的细胞中测量Akt和c-Jun N末端激酶（JNK）的活性，有或没有10添加 10 μM LY294002 后，Akt 活性降低。我们发现 LY294002 增加了 JNK 活性，已知 JNK 活性可介导 PNS 神经元中的某些类型的细胞死亡。我们还观察到 LY294002 在 HKIID1+JNKD1 +BDNF 中提高了 c-Jun 的表达。 PI3 激酶活性涉及小脑颗粒神经元中 Akt 活性的抑制和 JNK 活性的升高。 PI3-激酶-Akt 通路抑制 JNK 和 c-Jun 表达的激活，从而防止小脑颗粒神经元的神经元细胞死亡。

项目成果

山田雅司、畠中寛: "羊土社"「新アポトーシス実験法-基本操作と最先端の解析方-」培養中枢神経細胞を用いたアポトーシスの解析. 307(149-157) (1999)

Masashi Yamada、Hiroshi Hatanaka：“Yodosha”“新细胞凋亡实验方法 - 基本操作和尖端分析方法”使用培养的中枢神经细胞进行细胞凋亡分析 307（149-157）（1999）。

T. Satoh, T. Yamagata, Y. Ishikawa, M. Yamada, Y. Uchiyama and H. Hatanaka: "Regulation of reactive oxygen species by nerve growth factor but not by Bcl-2 as a novel mechanism of protection of PC12 cells from superoxide anion-induced death."J. Biochem. 12

T. Satoh、T. Yamagata、Y. Ishikawa、M. Yamada、Y. Uchiyama 和 H. Hatanaka：“通过神经生长因子而不是 Bcl-2 调节活性氧，作为保护 PC12 细胞免受侵害的新机制

下家浩二ら: "Inhibition of phosphatidylinositol 3-kinase activity elevates c-Jun-terminal kinase activity in apoptosis of cultured cerebellar granule neurons."Dev.Brain Res.,. 112,. 245-253 (1999)

Koji Shimoie 等人：“抑制磷脂酰肌醇 3-激酶活性可提高培养的小脑颗粒神经元细胞凋亡中的 c-Jun 末端激酶活性。”Dev.Brain Res., 112, 245-253 (1999)

稲村直子ら: "The role of p53 in DNA strand break-induced apoptosis in organotypic slice cultures from the mouse cerebellum."J.Neurosci.Res.,. (印刷中). (2000)

Naoko Inamura 等人：“p53 在小鼠小脑器官切片培养物中 DNA 链断裂诱导的细胞凋亡中的作用”。J.Neurosci.Res.（出版中）。

T. Araki, M. Yamada, H. Ohnishi, S. Sano, T. Uetsuki and H. Hatanaka: "Role of phosphatase activity of Shp-2 in BDNF signaling in cultured cerebral cortical neurons."J. Neurochem. 74. 659-668 (2000)

T. Araki、M. Yamada、H. Ohnishi、S. Sano、T. Uetsuki 和 H. Hatanaka：“Shp-2 磷酸酶活性在培养的大脑皮层神经元 BDNF 信号传导中的作用”。